A confocal laser microscope is configured such that laser light is focused on a specimen through an objective lens, a light flux of reflected light, scattered light, or fluorescent light generated on the specimen is transmitted by an optical system, and the light flux transmitted through a pinhole disposed at an optically conjugated position with respect to a light focusing point on the specimen is received on a detector. Disposing the pinhole makes it possible to filter the light generated on the specimen other than the light focusing point. Therefore, the confocal laser microscope is operable to acquire an image with a good S/N ratio.
Further, the confocal laser microscope is configured to acquire a planar image of a specimen by scanning the specimen with laser light along two directions (X-direction and Y-direction) orthogonal to each other, along a plane perpendicular to the optical axis. On the other hand, the confocal laser microscope is configured to acquire a plurality of tomographic images (Z-stack images) in Z-direction by changing the distance in the optical axis direction (Z-direction) between the objective lens and the specimen, whereby a three-dimensional image of the specimen is formed.
In observing a biospecimen, it is often the case that the biospecimen is observed through a cover glass in a state in which the biospecimen is immersed in a broth. Further, generally, the objective lens is designed so that an optimum imaging performance at a position immediately below the cover glass is best. In observing the inside of a biospecimen, it is necessary to acquire an image transmitted through a broth or biological tissues and having a certain depth at an observation position. Aberrations are generated in proportion to the distance from the position immediately below the cover glass to the observation position, and as a result, the resolution may be lowered.
Further, the cover glasses have variations in the thickness thereof within the tolerance range from the design value (e.g. 0.17 mm). Aberrations are generated in proportion to a difference between the actual thickness of the cover glass and the design thickness due to a difference between the refractive index (=1.525) of the cover glass and the refractive index (=1.38 to 1.39) of the biospecimen. Further, when the objective lens is an immersion lens, aberrations are generated in proportion to the depth of a biospecimen with respect to the observation position due to a difference between the refractive index of the biospecimen and the refractive index (=1.333) of water in the same manner as described above. As a result, the resolution to be obtained in observing a deep part of the biospecimen may be lowered.
As one means for solving the above defects, a correction ring has been proposed. The correction ring is a ring-shaped rotating member provided in an objective lens. The distances between lens groups constituting the objective lens is changed by rotating the correction ring. Aberrations due to an error in the thickness of the cover glass or observing a deep part of the biospecimen are cancelled by rotating the correction ring. A scale is marked on the correction ring. For instance, rough numerical values such as 0, 0.17, and 0.23 are indicated concerning the thickness of the cover glass. Adjusting the scale of the correction ring in accordance with the thickness of an actually used cover glass makes it possible to adjust the distances of the lenses in such a manner as to optimize the distances in accordance with the thickness of the cover glass (e.g. see Patent Literature 1).
Further, there is also known a technique of compensating for generated aberrations by a wave front conversion element. This technique is a matrix-drivable shape variable mirror element that is disposed on an optical path of a microscope, a wave front is modulated by the shape variable mirror element based on wave front conversion data measured in advance, and the modulated light wave is allowed to be incident on a specimen, whereby an aberration-corrected image with a high imaging performance is acquired (see e.g. Patent Literature 2).
As the wave front conversion element, a shape variable mirror element configured such that the shape of a reflection surface thereof is electrically controllable is used. When a plane wave is incident on the shape variable mirror element, and if the shape variable mirror element has a concave shape, the incident plane wave is converted into a concave wave front (the amplitude of a concave shape is doubled).